Several lignans, mostly new, were isolated from Larrea tridentata by assay-guided countercurrent chromatography (CCC). Using the secreted alkaline phosphatase bioassay of HIV Tat transactivation and the two- phase solvent systems composed of hexane:ethyl acetate: methanol:water, two major components (Gr and Lo) were identified as anti-HIV active principles. The chemical structure of the constituents of Gr (G-1 to G-4) and Lo (L-1 to L-4) were determined by GC/MS and NMR. After optimization of isolation conditions, a large-scale isolation with the chloroform: methanol:water system yielded five constituents (FB-1 to FB- 5). The most predominant anti-HIV compound FB-2 or 3prime-O-methyl nordihydroguaiaretic acid (3prime-O-methyl NDGA, denoted mal.4 (L-2) which occurs in 0.23% yield, was separated from its FB-1 isomer (4prime- O-methyl NDGA or L-1)(0.13% yield). These compounds may represent a new class of anti-HIV agents with important clinical relevance.